1. Field of the Invention
The present invention relates to media for use in electrophoresis and a method of using such media. More particularly the present invention relates to a medium for use in electrophoresis having N-methylurea and a gel based on polyacrylamide and a method involving the introduction of a sample to a medium, and applying an electric field across the medium.
2. Description of the Prior Art
Electrophoresis is a technique used for the separation of DNA, RNA, proteins and other macro- and micro-molecular compounds. Biomolecules, such as proteins, amino acids, peptides, nucleotides, and nucleic acids are charged or can be made to be charged such that they will move in solution when subjected to an electrical field. The velocity at which the biomolecules migrate in this electrical field depends on the size, electrical charge, and other physical properties of the molecules.
The medium for this electrophoretic migration is typically a gel slab or a gel matrix within a capillary or a channel. These gels are often polymer networks formed by the mixture of a polymer with a certain amount of a cross-linker. The amount of cross-linker used determines the pore size within the gel through which the biomolecules migrate. U.S. Pat. No. 5,055,517 describes such a cross-linked polymer gel.
The biomolecular components separate across the electrophoretic medium over time when the electrical field is applied across the medium. One method of detecting and identifying the separated components involves staining the biomolecules with a dye and comparing the migration times to known standards. Another method of detection and identification involves measuring the fluorescence of the separated components.
When the molecules are to be separated by their size or molecular weights a denaturing agent is often added to the gel medium to improve resolution of the separated components. Urea can be used as this denaturing agent. However, the advantages in resolution obtained with the use of urea are offset by a negative effect on background fluorescence.
The effect of urea and alkylureas, such as N-methylurea, on the thermal stability, structural properties, and preferential salvation changes accompanying the thermal unfolding of ribonuclease A is described in xe2x80x9cThermodynamic stability of ribonuclease A in alkylurea solutions and preferential salvation changes accompanying its thermal denaturation: A calorimetric and spectroscopic study,xe2x80x9d Protein Science 8:832-840 (1999). However, alkylureas have not previously been shown to improve the resolution of components separated by electrophoresis.
Accordingly, it would be desirable to provide a method, system, and a medium for electrophoresis that improves the resolution of components separated by electrophoresis while minimizing background fluorescence.
The present invention provides a medium for use in electrophoresis having N-methylurea and a gel capable of suspending the N-methylurea.
The present invention further provides an electrophoretic system having a medium and electrodes for introducing an electric field across the medium, wherein the medium has N-methylurea and a gel capable of suspending the N-methylurea.
The present invention further provides a method of electrophoretic separation involving the introduction of a sample volume to a medium, and applying an electric field across the medium, wherein the medium includes N-methylurea.